Fluorescent labelling negatively affects the physiology of Lactococcus lactis

Culturability on plates, growth behaviour in liquid media and acidification activity of three Lactococcus lactis strains was negatively affected by carboxyfluorescein diacetate (cFDA), the corresponding succinimidyl ester cFDA-SE, propidium iodide (PI) and TOTO-1 iodide. Single staining with the vitality dye cFDA-SE decreased the reproductive capability and acidification activity of L. lactis cells more than cFDA. Since TOTO-1 proved to have a higher toxicity than PI, the application of PI was favoured over TOTO-1 as counterstain for cells labelled with one of the vitality dyes. The overall extent to which double staining with cFDA/cFDA-SE and PI impaired bacterial physiology was determined by the dye with the greater influence on L. lactis cells during single staining. The observed strain-dependent differences in sensitivity highlight the importance of studying the impact of fluorescent labelling on cell physiology before using the dyes in combination with flow cytometric cell sorting for physiological characterisation of subpopulations.